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RNASeQC error while running example data

Hi all! I was trying to run a command from the example that was given on this webpage: http://www.broadinstitute.org/cancer/cga/rnaseqc_download They give example RNA-seq data to test the RNASeQC tool. I downloaded all files to the same directory, unpacked them and wanted to run the command: 

java -jar RNA-SeQC_v1.1.8.jar -n 1000 -s "TestId|ThousandReads.bam|TestDesc" -t gencode.v7.annotation_goodContig.gtf -r Homo_sapiens_assembly19.fasta -o ./testReport/ -strat gc -gc gencode.v7.gc.txt -BWArRNA human_all_rRNA.fasta -bwa ~/RNAseq/tools/bwa-0.7.15/bwa

I basically took their command and added the path to BWA and changed RNASeQC.jar to RNA-SeQC_v1.1.8.jar The .bam file is indexed. Unfortunatelly I didn't get anything but this:

RNA-SeQC v1.1.8.1 07/11/14
Retriving contig names from reference
         contig names in reference: 85
Loading GTF for Read Counting
Converting to refGene
Transcript objects to RefGen format:    3 s
Running IntronicExpressionReadBlock Walker ....
Arguments: [-T, IntronicExpressionReadBlock, --outfile_metrics, ./testReport//TestId/TestId.metrics.tmp.txt, -R, Homo_sapiens_assembly19.fasta, -I, ThousandReads.bam, -refseq, ./testReport//refGene.txt, -l, ERROR]

Could someone please tell me what I'm doing wrong? I'm very new to RNA-seq. It's probably something obvious but I can't find the mistake :(

rna-seq
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updated by GenoMax GenoMax • written by anjasta47 • anjasta47

Which version of java are you running?

• 1 views • link
written by GenoMax GenoMax

java version "1.7.0_71" OpenJDK Runtime Environment (rhel-2.5.3.1.el6-x86_64 u71-b14) OpenJDK 64-Bit Server VM (build 24.65-b04, mixed mode)

• 1 views • link
written by anjasta47 • anjasta47

I guess you did not print all of the error messages. I tried the test data out and I am getting the same error but in addition this 

org.broadinstitute.sting.utils.exceptions.UserException$MalformedBAM: SAM/BAM file ThousandReads.bam is malformed: SAM file doesn't have any read groups defined in the header.  The GATK no longer supports SAM files without read groups
at org.broadinstitute.sting.gatk.datasources.reads.SAMDataSource.<init>(SAMDataSource.java:276)
at org.broadinstitute.sting.gatk.GenomeAnalysisEngine.createReadsDataSource(GenomeAnalysisEngine.java:755)
at org.broadinstitute.sting.gatk.GenomeAnalysisEngine.initializeDataSources(GenomeAnalysisEngine.java:666)
at org.broadinstitute.sting.gatk.GenomeAnalysisEngine.execute(GenomeAnalysisEngine.java:227)
at org.broadinstitute.sting.gatk.CommandLineExecutable.execute(CommandLineExecutable.java:113)
at org.broadinstitute.sting.commandline.CommandLineProgram.start(CommandLineProgram.java:236)
at org.broadinstitute.sting.commandline.CommandLineProgram.start(CommandLineProgram.java:146)
at org.broadinstitute.cga.rnaseq.gatk.GATKTools.runIntronReadCount(GATKTools.java:226)
at org.broadinstitute.cga.rnaseq.ReadCountMetrics.runRegionCounting(ReadCountMetrics.java:244)
at org.broadinstitute.cga.rnaseq.ReadCountMetrics.runReadCountMetrics(ReadCountMetrics.java:59)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.runMetrics(RNASeqMetrics.java:225)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.execute(RNASeqMetrics.java:171)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.main(RNASeqMetrics.java:139)

Did you get this message as well?

• 1 views • link
written by GenoMax GenoMax

Thank you very much for your involvement! When I ran the the command the first time I got this message as well but when I ran it again I only got those few lines I posted.

• 1 views • link
written by anjasta47 • anjasta47

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